Download (0)

Full text




Dr. Mokshi Sharma


, Dr. G K Sangha



Department of Zoology, Punjab Agricultural University (India)


Pesticide hazards have been accentuated by the sharp rise in their agricultural, industrial and domestic use. The extensive use of dimethoate poses a health hazard to animals and humans because of its persistence in soil and crops. Our study investigated the toxic effects of the dimethoate at different dose level, an organophosphorous pesticide, on organ like liver of albino rat. Dimethoate was administered orally for one month at doses 1/20th, 1/40th and 1/80th on the basis of LD50. Half of the experimental animals were sacrificed and the rest were left to study the inbuilt capacity of the animals to metabolize the toxicant. Treated mice were showed many histopathological alterations in liver like, vacuolation, blood congestion, and high lymphocytic infiltration around the central vein, enlargement of hepatic sinusoids, hepatocellular damage, necrosis, and increase in number of kuffer cells, lessions and hemorrhage. However, these alterations were recovered in the next set of experimental animals showing the inbuilt capacity of the organism to metabolize the toxicant.


Dimethoate, Histopathology, Albino rat.

Short Profile



The environmental pollution is one of the most serious problems that faces mankind in this century. There are

many types of pollutants that interfere with our-life both directly and indirectly. Furthermore, potential future

hazards to human health and wildlife can be created by residues from some long-lived pesticides that may build

up in the food chain and cause widespread contamination of the environment (El-Sebae, 1993; Zaahkouk et al., 2000). Nowadays, the hazards of pesticides have been accentuated by the sharp rise in their use by farmers,

industrialists and householders alike. Pesticide poisoning resulting in death occurs mainly in developing

countries where the consumption of these products is high (Banerjee et al., 1999). Organophosphate (OP)

compounds are the most toxic pesticides causing environmental pollution and potential health hazards (Agdi et

aI., 2000). Being biologically active compounds, pesticides have been thoroughly tested for safety and usefulness prior to their agricultural use. However their misuse may cause drastic effects to humans, and the

environment (GIFAP, 1984).

Dimethoate, are the widely used systemic pesticides in agriculture against a wide range of insects, mites and

fungal diseases of fruits, vegetables, or namental plants and field crops as both systemic and contact pesticides

and are also used indoor to control houseflies (Meister, 1992; Farag et al., 2006; Brkic et al., 2008 and Muthuvivegandave et al., 2011). The irrational and excessive use of these pesticides plays a crucial role in the


chemicals and drugs. Hepatotoxicity is therefore an important end point in the evaluation of the effect of a

particular xenobiotic. Clinical chemistry and histopathological evaluations are commonly used methods for

detecting organ specific effects related to chemical exposure (Travlos et al., 1996; Crissmann et al., 2004). Dimethoate has acute oral toxicity of 150mg/kg by in mice and has been classified as moderately hazardous

(IPCS/WHO., 2001; Kidd, et al., 1991). Also, Poet et al., 2003, studied the effect of two organophosphorous pesticide on the hepatic and intestinal metabolism in the rat and found that the metabolizing enzymes

responsible for both the bioactivities and detoxification are present in the small intestine at lower level than the

liver but still significant. Also, Bulusu and Chakravartylndira et al., 1992, studied nucleic acid and protein

propel in normal and malnourished rat liver on exposure to organophosphorous group of pesticides and found

that were affected in both. Moreover it has been proven to accumulate in the testis where it persisted for weeks

even after oral administration was stopped (Afifi et al., 1991) and inhabited steroidogenesis primarily by

blocking transcription of the steroidogenic acute regulatory (STAR) gene (Walsh 2000). Few studies have been

made on the histopathological effects of dimethoate (Thangavel et al., 1994; Persis et al., 2001). The present studies have been aimed to investigate the histopathological effects of the dimethoate (an organophosphorous

pesticide) that is widely used in some agricultural areas in India on some organ of albino rat.



2.1 Chemicals and Plant extract

All chemicals were purchased from SDFCL (SD Fine-Chem Ltd) and SRL (Sissco Research Laboratories Pvt

Ltd). All chemical used were either of analytical grade or the highest purity commercially available. Standard

rat feed was purchased from Ashirwad Industries, Mohali, India.

2.2 Animals

The female albino rats aging 9-12 weeks and weighing 140-170 grams were procured from the Department of

Livestock Production and Management, GADVASU, Ludhiana and were maintained in laboratory conditions

with standard pelleted rat feed and water provided ad libitum.The animals were kept in a room in which the humidity and temperature were environmentally controlled. All methods and procedures of animal handling

during research were conducted in accordance with the guidelines of Committee for the Purpose of Control and

Supervision of Experiments on Animals (CPCSEA), India and experiments conducted in the present study were

duly approved by Institutional Animal Ethics Committee (IAEC), Guru Angad Dev Veterinary and Animal

Sciences University, Ludhiana vide letter no IAEC/2014/125-53 dated 13/08/2014.

2.3 Experimental design

Female albino rats were acclimatized for ten days in laboratory conditions and were divided into eight groups

with six rats in each group.

Group I (Control) animals received feed + distilled water, olive oil and served as control

Group II 1/20th of dimethoate by oral intubation

Group III 1/40th of dimethoate by oral intubation


GroupV Olive oilfor15 days and left without any treatment for next 15 days then sacrificed.

GroupVI 1/20th of dimethoate for15 days and left without any treatment for next 15 days then sacrificed.

Group VII 1/40th of dimethoate for15 days and left without any treatment for next 15 days then sacrificed.

Group VIII 1/80th of dimethoate for15 days and left without any treatment for next 15 days then


At the end of 15 days, mice from exposed Groups (I, II, III & IV) were anesthetize and dissected, liver was

removed and fixed for histopathological investigations using 10% neutral formal saline. Tissue was processed

by routine histological techniques, sectioned at 6μm, stained with hematoxylin and eosin. Finally stained

sections were examined under the high magnifying microscope and subsequently micrographs were taken. At

the end of experimental period, rat from (Group V, VI, VII &VIII) were taken in batches dissected and

examined for histopathological investigations. O


3.1 Histopathological Study of Liver:

Sections from the liver were prepared from both control and treated mice and were examined under the high

magnifying light microscope. The microscopic observations were reported and the intensity of changes was

tabulated in Table 1, and presented by the Figs. 1. These changes include the presence of more endothelial cells

scattered among the hepatocytes and some vacuolation in the liver texture (Fig A to F). Some sections of liver

from mice of treatment groups (I to III) treated with dimethoate pesticides showed rupture in some hepatocytes,

many other sections of the treatment sets showed dense lymphocytic infiltration round the central vein and dark

stained hepatocytic nuclei indicating cell pycnosis. A number of hepatocytes were found to contain shrinked

nuclei, while the liver sections belonging to the metabolized groups showed minor changes represented by

group V to VIII. LI


Table-I Histopathological Changes observed in Liver




1 Congestion + ++ ++ + + ++ + +

2 Cell rupture No Mild Moderate Moderate No Moderate No No

3 Vaculation + +++ ++ ++ No + No No

4 Lymphocytic


Occasional ++ ++ + No ++ + +

5 Nuclear death No ++ + + No No No No

6 Hemorrhage No ++ + + No periportal

round cell


No No

7 Enlargement of

control & portal



8 Hepatocellular


No ++ ++ + No Perivescular




No No

9 Increase in the


of kuffer cells

No +++ ++ + No mild No No

10 Necrosis No ++ + + No Focal area

show necrosis

No No

11 Subcapsular

stimulation of



No ++ + + No No No No

12 Enlargement of



No +++ ++ ++ No Mild No No

13 Parenchymatous

cells showing


vacuolization &

degeneration in


No ++ + + No No No No

14 Hepatocytes whic h

may show variation


lipid Like bodies of

various sizes

Normal +++ ++ + No Mild No No



The animal model used in this study was used to assess the adverse effects of pesticides on laboratory animals

(Costa et al., 1989). While, investigating the histopathological effects of dimethoate pesticides on Liver of

albino rat, the results showed mild to severe effects on the organ studied. Some significant results were found

while investigating the histopathological parameters. The Histology section of Liver showed to be more severely

affected by all the doses of dimethoate. The changes reported include lymphocytic infiltration, congestion,

nuclear death,enlargement of hepatic sinusoids, hepatocellular damage, parenchymatous cells showing

cytoplasmic vacuolation and degeneration in nuclei. To a bit extent, cell rupture, increase in kuffer cell

number,hemorrhage and some hepatocytes which showed variation in lipid like bodies of various sizes were

also observed. The role of liver in xenobiotic metabolism is very essential as it contains a large number of

enzymes responsible for xenobiotic metabolism. Accordingly as per its role it is more susceptible to chemical

injury (Sharkoori, Alam & Ali., 1990). Section through the liver showed chronic inflammation,congestion

which could lead to jaundice and death of hepatocyte (necrosis).In an earlier study, Khogali et al. (2005),

reported Dimethoate-induced hepatic pycnosis,vacuolation, blood congestion and high lymphatic infiltration

around the central vein; Thangevel et al., 1994 reported the toxic effect of an organophosphate pesticide dimecron and acarbamate fungicide cumin on the histology of liver showed the destruction of hepatocytes and

protrusion of nuclei. While oral administration of Carbosulfan at 48 mg/kg/day for 5, 10, 20 and 30 days in

albino mice resulted in dilation of central vein and sinusoids between hypertrophied hepatocytes,vacuolization

and hyalinization of hepatocytes with loss of radial arrangement, suggesting that theinsecticide had adverse

effects on liver functions leading to histological and physiological impairments (Ksheerasagar and Kaliwal,

2006). The lymphocytic infiltration observed in some controlled mice might be due to the groundnut oil or due

to the hygienic condition other than the chemical effects. Liver tissues from mice treated with high doses of the

dimethoate showed hepatocytic nuclear death or cell pycnosis. Similar effects were reported by Singh et al.,

(1994). The evidences of liver damage like cord disarray hypertrophy and disintegration of hepatocytes showing

different sizes of nuclei, lymphocytic infiltration, in addition to sinusoidal blood congestion and hemorrhage

was also reported by many investigators using various chemicals and toxicants with different animals including

fishes (Persis and Kalairasi, 2001; Abdu Rabou.,1996; Shakoori., 1992; Khogali 1989).Recently, El-bendary et

al. (2014) evaluated histopathological abnormalities associated with Profenifos and Chlorpyrifos exposure in

male albino mice where liver showed hepatic cell damage with degenerative changes including congestion of

blood vessels, vacuolar degeneration of hepatic cells, focal infiltration and mononuclear cells, dilated central

vein and other hepatic blood vessels, necrosis of hepatic cells, disorganized with the formation of denoid

structure and hepatocytomelagy; whereas kidney showed hemorrhage, oedema, necrosis and glomeruli

shrinkage. Since, the present feed treatments of albino rat with Dimithoate clearly demonstrated significantly

changes in the histopathology of hepatic tissue, it could be possible that prolonged exposure to this insecticide in

man may play a significant role in aggravating such diseases as chronic liver failure. In addition, structural

changes to hepatic tissue such as haemorrhage, congestion, vacuolation and erosion may also lead to acute liver

damage. The present results could thus be exploited as a potential biomarker of common insecticide toxicity in



The present findings clearly demonstrate that Dimithoate is capable of inducing dosedependent morphometric

i.e. histopathological change in the liver of the exposed rat. According to these data, it is suggested that systemic

insecticide like Dimethoate exposure might cause hazardous effects, especially at high doses, to man and

environment. For field and domestic uses of this insecticide, quantities and mode of usage need to be strictly

monitored to minimize the possibility of its exposure to non-target organisms including human beings. This can

be achieved through public health education to make people aware of the hazardous effects of this chemical.

Due attention also is to be paid for a delayed period of field application of this insecticide to avoid its possible

adverse effects to consumers, who should be warned of the potential risk of Dimithoate contamination of food

and drinking water in the country.


1. A. N. Abdu Rabou. On the Hazards of Pesticide Use and Misuse in Khartoum State and Some Histopathological

Studies on the Effects of Carbamates on Experimental Animals. MSc Thesis in Environmental Studies. Sudan:

University of Khartoum, 1996.

2. N. A. Afifi, A. Ramadan, M. I. Abd El-Aziz and E. E. Saki. Influence of dimethoate on testicular and epididymal

organs, testosterone plasma level and their tissue residues in rats, Dtsch tierärztl Wschr 98(2),1991, 419–423.

3. K. Agdi, A. Bouaid, A. M. Esteban, P. F. Hernando, A. Azmani, and C. Camara Removal of atrazine and four

organophosphorus pesticides from environmental waters by diatomaceous earth remediation method. Journal of

Environmental Monitoring 2(5), 2000, 420–423.

4. M. Al-Haj, A. Nasser and A. Anis. Survey of pesticides used in qat cultivation in dhale and yafe and their

adverse effects. Journal of National Applied Science 9(1), 2005, 103-110.

5. B. D. Banerjee, V. Seth, Bhattacharya A, Pasha ST, and Chakraborty AK (1999) Biochemical effects of some

pesticides on lipid peroxidation and free-radical scavengers. Toxicology Letters 107(1-3): 33–47.

6. D. V Brkic, S. L. Vitorovic, S. M. Gasic and N. K. Neskovic. Carbofuran in water:subchronic toxicity to rats.

Environmental Toxicology and Pharmacology, 25, 2008, 334-341.

7. S. Bulusu and I. Chakravarty. „Nucleic Acid and Protein Profile in Normal and Malnourished Rat Liver on

Exposure to Organophosphorous Group of Pesticides.” International Journal of Tounco. Occupl Environment

Hlt., 1(2), 1992, 46-51.

8. L. G. Costa, G. Olibet, D. Wu and S. D. Murphy. Acute and chronic effects of the pesticide amitraz on a 2-

adrenoceptors in the mouse brain. Toxicological Letters. 47(2), 1989, 135-143.

9. J. W. Crissman, D. G. Goodman, P. K. Hildebrandt, R. R. Maronpot, D. A. Prater and J. H. Riley et al., Best

practice guideline: Toxicologic histopathology, Toxicology Pathol 32, 2004, 126–131.

10. H. M. El-bendary, M. H. Shaker, A. A. Saleh, S. E. Negm, M. E. Khadey, F. A. H. Elden. Histopathological

changes associated with exposure of male mice to prefenofos and chlorpyrifos. Ann. Res. Rev. Biol., 4(5),

2014,766-777. www.sciencedomain.org

11. A. H. El-Sebae, M. Abou-Zeid and M. Saleh. Status and environmental impact of toxaphene in the third world-a

case study of African Agriculture Chemosphere, 27(10),1993, 2063-2072.

12. A. T. Farag, T. A. Karkour and A. El-Okazy. Developmental toxicity of orally administered technical dimethoate


13. P. H. Howard, (Ed.). Handbook of Environmental Fate and Exposure Data for Organic Chemicals.Chelsea: Lewis

Publishers, Vol. 3, 1991.

14. IPCS/WHO. Classification of pesticides by hazard and guidelines to classification. Switzerland:Geneva; 2001.

15. F. A. Khogali. and S. A. Gumaa. “Some Histopathological Effects of Lethal and Sublethal Doses of 2,4-D on

Oreochromis niloticus Linn. and Tilapia zillii Grev. from the White Nile near Khartoum.” Sudan Journal of

Science, 4 1989, 86-99.

16. F. A. Khogali, J. B. Sheikh, S. A. Rahman, A. A. Rahman, M. H. Daghestani. Histopathological and

hematological effects of dimethoate 40EC on some organs of albino mice. Journal King Saud University, 18 Sci.

(2), 2005, 73-87.

17. H. Kidd and D. R. James, Editors. The agrochemicals handbook (3rd ed.), Royal Society of Chemistry

Information Services, Cambridge, UK (1991).

18. R. L. Ksheerasagar and B. B. Kaliwal. Histological and biochemical changes in the liver of albino mice on

exposure to insecticide, Carbofuran. Caspian Journal Environment Science, 4(1), 2006, 67-70.

19. R. T. Meister. Farm chemicals handbook. Willoughby, OH: Meister Publishing Company. 1992

20. V. Muthuviveganandave, P. Muthurman, I. Hawang and K. Srikumar. Biochemical and Histopathological

changes induced by low doses of carbendazim on testis of male albino rat. International Journal of Pharmacy and

Biological Sciences, 1(4), 2011, 572-576.

21 V. T. Persis and J. M. V Kalaiarasi. “Histopathological Responses of Mystus vittatus to Chronic Sublethal and Acute Lethal Toxicity of an Organophosphate Pesticide.” Journal of Expt Zoo India, 4(1), 2001,103-108.

22. T. S Poet, Wu. H, A. A. Kousba, and C. Timehalk. “In vitro Rat Hepatic and Intestintnal Metabolism of the Organophosphate Pesticides Chlorpyrifos and Diazinon.” Toxicological Sciences, 72, 2003, 193-200.

23. A. R. Shakoori, J. Alam , F. Aziz , F. Aslam and M. Sabir. “Toxic Effects of Bifenthrin (Talstar) on the Liver of

Gallus domesticu‟‟. Journal Ecotoxicology Environment Monit., 2(1), 1992, 1-11.

24. A. R. Sharkoori, J. Alam and S. S. Ali, “Toxic Effects of Talstar: A New Synthetic Pyrethroid, on Blood and Liver

of Rabbit.” Pakistan Journal of Zoology, 22(3), 1990, 289-300.

25. N. Singh and A. K Srivastava. “Formothion Induced Haematological Changes in the Fresh Water Indian Catfish

Meteropneustes fossili”. Journal of Ecotorico Environ. Monitor, 4(2), 1994, 137-140.

26. P. Thangavel, I. Baby Skaikila and M. Ramaswamy. „„Synergestic Effect of Dimecroncuman L. on the Histology

of Liver and Brain of Sarotherodon mossambicus‟‟. Jecobio, 6(3), 1994, 173-178.

27. G. S. Travlos, R. W. Morris, M. R. Elwell, A. Duke, S. Rosenblum and M.B. Thompson, Frequency and

relationship of clinical chemistry and liver and kidney histopathology findings in 13-week toxicity studies in rats,

Toxicology 107, (1996), 17–29.

28. L. P. Walsh, D. R. Webster and D.M. Stocco. Dimethoate inhibits steroidogenesis by disrupting transcription of

steroidogenic acute regulatory (StAR) gene, Journal of Endocrinology 167 (2000), 253–263.

29. S. A. M. Zaahkouk, E. G. E. Helal, T. E. I. Abd-Rabo and S. Z .A. Rashed,. Carbamate toxicity and protective

effect of vit. A and vit. E on some biochemical aspects of male albino rats. The Egyptian Journal of Hospital


Figure 1 Showing Liver sections from different treatment groups of Albino rat

Figure 1

Showing Liver sections from different treatment groups of Albino rat p.4